Abstract
Zinc transporter 2 (ZnT-2) is one of the cellular factors responsible for Zn homeostasis. Upon Zn overload, ZnT-2 transports Zn into excretory vesicle to reduce cellular Zn. We investigated in this study the molecular mechanism involved in the regulation of human ZnT-2 (hZnT2) gene. When treated with Zn, hZnT-2 gene expression increased in a dose- and time-dependent manner. Besides to Zn, Cd is also a potent inducer of the gene. Metal response element (MRE)-binding transcription factor 1 (MTF-1) regulated hZnT-2 expression since over-expression of MTF-1 increased the transcription whereas knock-down of MTF-1 reduced the expression. Analysis of the hZnT-2 promoter region revealed that five putative MREs are present. A series deletion of the promoter region was conducted to investigate the MREs required for Zn induction. The result showed that at least two MREs were required. Mutation for each MRE was also performed to evaluate the significance of individual MRE for Zn induction. Surprisingly, only one MRE (MREb) is sufficient for the metal induction. Besides to MREs, a sequence for zinc-finger E-box binding homeobox (ZEB) binding is present next to MREc. Mutation or deletion of the ZEB biunding site resulted in an increase of the basal and induced hZnT-2 promoter activity. Knock-down of ZEB-1 increased the expression of hZnT-2, and over-expression ZEB-2 reduced hZnT-2 expression. However, over-expression ZEB-1 or knock-down of ZEB-2 did not affect hZnT-2 expression. Results from this study show that MTF-1 stimulates but ZEB represses the expression of hZnT-2 under Zn overload.